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Biochem Biophys Res Commun. 2007 Oct 26;362(3):646-50. Epub 2007 Aug 13.

Genetic design of conditional D-glutamate auxotrophy for Bacillus subtilis: use of a vector-borne poly-gamma-glutamate synthetic system.

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  • 1Department of Bioresources Science, Kochi University, Nankoku, Kochi 783-8502, Japan. ashiuchi@cc.kochi-u.ac.jp

Abstract

Bacillus subtilis possesses two glutamate racemase isozymes, RacE and YrpC. For the first time, we succeeded in constructing glutamate racemase-gene disruptants of B. subtilis. Phenotypic analysis of their D-glutamate auxotrophy indicated that the RacE-type glutamate racemase is important for ensuring maximum growth rate but dispensable. The YrpC-type glutamate racemase probably operates as an anaplerotic enzyme for RacE, especially under liquid culture conditions. We found novel applicability of RacE-less mutants inheriting only a marginal activity for endogenous D-glutamate supply, viz. the employment for the in vivo identification of D-glutamate-consuming systems. In fact, the genetic induction of a poly-gamma-glutamate synthetic system led a RacE-less mutant to severe growth suppression, which was overcome in the presence of a high concentration of exogenous D-glutamate. The results indicate that a significant amount of D-glutamate is consumed during poly-glutamate biosynthesis. To our knowledge, this is the first report of conditional D-glutamate auxotrophy for B. subtilis.

PMID:
17720142
[PubMed - indexed for MEDLINE]
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